The International Society for Vaccines is an organization that engages, supports, and sustains the professional goals of a diverse membership in all areas relevant to vaccines - 2018 ISV Annual Congress

Generation of influenza A viruses as live but replication-incompetent virus vaccines

Science. 2016. 354:1170-1173

Authors:

Longlong Si, Huan Xu, et al. 2016

Abstract:

The conversion of life-threatening viruses into live but avirulent vaccines represents a revolution in vaccinology. In a proof-of-principle study, we expanded the genetic code of the genome of influenza A virus via a transgenic cell line containing orthogonal translation machinery. This generated premature termination codon (PTC)–harboring viruses that exerted full infectivity but were replication-incompetent in conventional cells. Genome-wide optimization of the sites for incorporation of multiple PTCs resulted in highly reproductive and genetically stable progeny viruses in transgenic cells. In mouse, ferret, and guinea pig models, vaccination with PTC viruses elicited robust humoral, mucosal, and T cell–mediated immunity against antigenically distinct influenza viruses and even neutralized existing infecting strains. The methods presented here may become a general approach for generating live virus vaccines that can be adapted to almost any virus.

The differentiation of human memory CD8 T cells is not well understood. Here we address this issue using the live yellow fever virus (YFV) vaccine, which induces long-term immunity in humans. We used in vivo deuterium labelling to mark CD8 T cells that proliferated in response to the virus and then assessed cellular turnover and longevity by quantifying deuterium dilution kinetics in YFV-specific CD8 T cells using mass spectrometry. This longitudinal analysis showed that the memory pool originates from CD8 T cells that divided extensively during the first two weeks after infection and is maintained by quiescent cells that divide less than once every year (doubling time of over 450 days). Although these long-lived YFV-specific memory CD8 T cells did not express effector molecules, their epigenetic landscape resembled that of effector CD8 T cells. This open chromatin profile at effector genes was maintained in memory CD8 T cells isolated even a decade after vaccination, indicating that these cells retain an epigenetic fingerprint of their effector history and remain poised to respond rapidly upon re-exposure to the pathogen.

rVSV-ZEBOV is a recombinant, replication competent vesicular stomatitis virus-based candidate vaccine expressing a surface glycoprotein of Zaire Ebolavirus. We tested the effect of rVSV-ZEBOV in preventing Ebola virus disease in contacts and contacts of contacts of recently confirmed cases in Guinea, west Africa.

Methods:

We did an open-label, cluster-randomised ring vaccination trial (Ebola ça Suffit!) in the communities of Conakry and eight surrounding prefectures in the Basse-Guinée region of Guinea, and in Tomkolili and Bombali in Sierra Leone. We assessed the efficacy of a single intramuscular dose of rVSV-ZEBOV (2×107 plaque-forming units administered in the deltoid muscle) in the prevention of laboratory confirmed Ebola virus disease. After confirmation of a case of Ebola virus disease, we definitively enumerated on a list a ring (cluster) of all their contacts and contacts of contacts including named contacts and contacts of contacts who were absent at the time of the trial team visit. The list was archived, then we randomly assigned clusters (1:1) to either immediate vaccination or delayed vaccination (21 days later) of all eligible individuals (eg, those aged ≥18 years and not pregnant, breastfeeding, or severely ill). An independent statistician generated the assignment sequence using block randomisation with randomly varying blocks, stratified by location (urban vs rural) and size of rings (≤20 individuals vs >20 individuals). Ebola response teams and laboratory workers were unaware of assignments. After a recommendation by an independent data and safety monitoring board, randomisation was stopped and immediate vaccination was also offered to children aged 6-17 years and all identified rings. The prespecified primary outcome was a laboratory confirmed case of Ebola virus disease with onset 10 days or more from randomisation. The primary analysis compared the incidence of Ebola virus disease in eligible and vaccinated individuals assigned to immediate vaccination versus eligible contacts and contacts of contacts assigned to delayed vaccination. This trial is registered with the Pan African Clinical Trials Registry, number PACTR201503001057193.

Findings:

In the randomised part of the trial we identified 4539 contacts and contacts of contacts in 51 clusters randomly assigned to immediate vaccination (of whom 3232 were eligible, 2151 consented, and 2119 were immediately vaccinated) and 4557 contacts and contacts of contacts in 47 clusters randomly assigned to delayed vaccination (of whom 3096 were eligible, 2539 consented, and 2041 were vaccinated 21 days after randomisation). No cases of Ebola virus disease occurred 10 days or more after randomisation among randomly assigned contacts and contacts of contacts vaccinated in immediate clusters versus 16 cases (7 clusters affected) among all eligible individuals in delayed clusters. Vaccine efficacy was 100% (95% CI 68·9-100·0, p=0·0045), and the calculated intraclass correlation coefficient was 0·035. Additionally, we defined 19 non-randomised clusters in which we enumerated 2745 contacts and contacts of contacts, 2006 of whom were eligible and 1677 were immediately vaccinated, including 194 children. The evidence from all 117 clusters showed that no cases of Ebola virus disease occurred 10 days or more after randomisation among all immediately vaccinated contacts and contacts of contacts versus 23 cases (11 clusters affected) among all eligible contacts and contacts of contacts in delayed plus all eligible contacts and contacts of contacts never vaccinated in immediate clusters. The estimated vaccine efficacy here was 100% (95% CI 79·3-100·0, p=0·0033). 52% of contacts and contacts of contacts assigned to immediate vaccination and in non-randomised clusters received the vaccine immediately; vaccination protected both vaccinated and unvaccinated people in those clusters. 5837 individuals in total received the vaccine (5643 adults and 194 children), and all vaccinees were followed up for 84 days. 3149 (53·9%) of 5837 individuals reported at least one adverse event in the 14 days after vaccination; these were typically mild (87·5% of all 7211 adverse events). Headache (1832 [25·4%]), fatigue (1361 [18·9%]), and muscle pain (942 [13·1%]) were the most commonly reported adverse events in this period across all age groups. 80 serious adverse events were identified, of which two were judged to be related to vaccination (one febrile reaction and one anaphylaxis) and one possibly related (influenza-like illness); all three recovered without sequelae.

Interpretation:

The results add weight to the interim assessment that rVSV-ZEBOV offers substantial protection against Ebola virus disease, with no cases among vaccinated individuals from day 10 after vaccination in both randomised and non-randomised clusters.

An estimated 100 million people have symptomatic dengue infection every year. This is the first report of a phase 3 vaccine efficacy trial of a candidate dengue vaccine. We aimed to assess the efficacy of the CYD dengue vaccine against symptomatic, virologically confirmed dengue in children.

Methods:

We did an observer-masked, randomised controlled, multicentre, phase 3 trial in five countries in the Asia-Pacific region. Between June 3, and Dec 1, 2011, healthy children aged 2–14 years were randomly assigned (2:1), by computer-generated permuted blocks of six with an interactive voice or web response system, to receive three injections of a recombinant, live, attenuated, tetravalent dengue vaccine (CYD-TDV), or placebo, at months 0, 6, and 12. Randomisation was stratified by age and site. Participants were followed up until month 25. Trial staff responsible for the preparation and administration of injections were unmasked to group allocation, but were not included in the follow-up of the participants; allocation was concealed from the study sponsor, investigators, and parents and guardians. Our primary objective was to assess protective efficacy against symptomatic, virologically confirmed dengue, irrespective of disease severity or serotype, that took place more than 28 days after the third injection. The primary endpoint was for the lower bound of the 95% CI of vaccine efficacy to be greater than 25%. Analysis was by intention to treat and per procotol. This trial is registered with ClinicalTrials.gov, number NCT01373281.

Findings:

We randomly assigned 10 275 children to receive either vaccine (n=6851) or placebo (n=3424), of whom 6710 (98%) and 3350 (98%), respectively, were included in the primary analysis. 250 cases of virologically confirmed dengue took place more than 28 days after the third injection (117 [47%] in the vaccine group and 133 [53%] in the control group). The primary endpoint was achieved with 56·5% (95% CI 43·8–66·4) efficacy. We recorded 647 serious adverse events (402 [62%] in the vaccine group and 245 [38%] in the control group). 54 (1%) children in the vaccine group and 33 (1%) of those in the control group had serious adverse events that happened within 28 days of vaccination. Serious adverse events were consistent with medical disorders in this age group and were mainly infections and injuries.

Interpretation:

Our findings show that dengue vaccine is efficacious when given as three injections at months 0, 6, and 12 to children aged 2–14 years in endemic areas in Asia, and has a good safety profile. Vaccination could reduce the incidence of symptomatic infection and hospital admission and has the potential to provide an important public health benefit.

Respiratory syncytial virus (RSV) is the leading cause of hospitalization for children under 5 years of age. We sought to engineer a viral antigen that provides greater protection than currently available vaccines and focused on antigenic site Ø, a metastable site specific to the prefusion state of the RSV fusion (F) glycoprotein, as this site is targeted by extremely potent RSV-neutralizing antibodies. Structure-based design yielded stabilized versions of RSV F that maintained antigenic site Ø when exposed to extremes of pH, osmolality, and temperature. Six RSV F crystal structures provided atomic-level data on how introduced cysteine residues and filled hydrophobic cavities improved stability. Immunization with site Ø–stabilized variants of RSV F in mice and macaques elicited levels of RSV-specific neutralizing activity many times the protective threshold.

In the RV144 trial, the estimated efficacy of a vaccine regimen against human immunodeficiency virus type 1 (HIV-1) was 31.2%. We performed a case-control analysis to identify antibody and cellular immune correlates of infection risk.

Methods:

In pilot studies conducted with RV144 blood samples, 17 antibody or cellular assays met prespecified criteria, of which 6 were chosen for primary analysis to determine the roles of T-cell, IgG antibody, and IgA antibody responses in the modulation of infection risk. Assays were performed on samples from 41 vaccinees who became infected and 205 uninfected vaccinees, obtained 2 weeks after final immunization, to evaluate whether immune-response variables predicted HIV-1 infection through 42 months of follow-up.

Results:

Of six primary variables, two correlated significantly with infection risk: the binding of IgG antibodies to variable regions 1 and 2 (V1V2) of HIV-1 envelope proteins (Env) correlated inversely with the rate of HIV-1 infection (estimated odds ratio, 0.57 per 1-SD increase; P=0.02; q=0.08), and the binding of plasma IgA antibodies to Env correlated directly with the rate of infection (estimated odds ratio, 1.54 per 1-SD increase; P=0.03; q=0.08). Neither low levels of V1V2 antibodies nor high levels of Env-specific IgA antibodies were associated with higher rates of infection than were found in the placebo group. Secondary analyses suggested that Env-specific IgA antibodies may mitigate the effects of potentially protective antibodies.

Conclusions:

This immune-correlates study generated the hypotheses that V1V2 antibodies may have contributed to protection against HIV-1 infection, whereas high levels of Env-specific IgA antibodies may have mitigated the effects of protective antibodies. Vaccines that are designed to induce higher levels of V1V2 antibodies and lower levels of Env-specific IgA antibodies than are induced by the RV144 vaccine may have improved efficacy against HIV-1 infection.

We used a systems biological approach to study innate and adaptive responses to influenza vaccination in humans, during 3 consecutive influenza seasons. Healthy adults were vaccinated with inactivated (TIV) or live attenuated (LAIV) influenza vaccines. TIV induced greater antibody titers and enhanced numbers of plasmablasts than LAIV. In TIV vaccinees, early molecular signatures correlated with, and accurately predicted, later antibody titers in two independent trials. Interestingly, the expression of Calcium/calmodulin-dependent kinase IV (CamkIV) at day 3 was inversely correlated with later antibody titers. Vaccination of CamkIV −/− mice with TIV induced enhanced antigen-specific antibody titers, demonstrating an unappreciated role for CaMKIV in the regulation of antibody responses. Thus systems approaches can predict immunogenicity, and reveal new mechanistic insights about vaccines.

ISV Nomination Statement:

A major challenge in vaccinology is that the effectiveness of vaccination can only be ascertained after vaccinated individuals have been exposed to infection. In 2009, Bali Pulendran and colleagues pioneered the use of a systems biological approach to study the global picture of the immune response to one of the most successful human vaccines ever developed, the live attenuated vaccine against yellow fever (Nat Immunol. 2009 Jan;10(1):116-25). Using this approach the investigators were able to identify signatures of gene expression in the blood of healthy humans, a few days after vaccination that could predict with up to 90 percent accuracy the strength of the immune response, weeks or months after yellow fever vaccination. In this ISV Paper of the Year (Nat Immunol. 2011 Jul 10;12(8):786-95), the same group, in collaboration with the group of Rafi Ahmed (an ISV member) and colleagues, now extend this approach to the seasonal influenza vaccines over the course of three influenza seasons. By studying gene expression patterns in the blood a few days after vaccination, the investigators were able to identify “signatures” that were capable of predicting the magnitude of the later immune response, with >90% accuracy. Importantly one of the genes in the signature, CAMK4 whose expression was negatively correlated with antibody titers, revealed an unappreciated role for CAMK4 in B cell responses. This landmark study, together with the previous work from the same group, demonstrates the use of systems biological approaches in predicting vaccine efficacy, and highlights one of the ways for the future of vaccinology - use of systems biology tools to perform sophisticated human studies that gives specific hypothesis to be tested experimentally